ABSTRACT Hematopoietic stem cells (HSCs) maintain tissue homeostasis and replenish blood system upon stresses. HSCs have evolved unique mechanisms to maintain genome and proteome integrity throughout life. While the genome integrity safeguard mechanisms have been extensively studied, little is known about how proteome integrity is maintained in HSCs and how dormant HSCs are protected from protein damage during development and physiological stresses. This proposal will address this knowledge gap. We recently found that the Sel1L ER-associated degradation (ERAD) pathway plays an essential role in the maintenance of HSCs. ERAD is the principal protein quality control mechanism responsible for targeting misfolded proteins in the ER for cytosolic proteasomal degradation. The Sel1L-Hrd1 complex is the most conserved branch of ERAD. Using a conditional knockout mouse model, we found that Sel1L deletion in hematopoietic cells significantly reduced steady-state HSC frequency and led to complete loss of HSC reconstitution capacity in stress conditions including bone marrow transplantation and 5-?uorouracil (5-FU) mediated myeloablation. Interestingly, Sel1L deletion did not induce apoptosis or impair HSC engraftment. In contrast, we observed increased HSC cycling and reduced numbers of quiescent HSCs in Sel1L knockout mice. These data demonstrate the critical function of Sel1L ERAD in HSC maintenance and suggest a novel role for ER protein quality control machinery in regulating stem cell quiescence and self-renewal. ERAD monitors and regulates the maturation of transmembrane proteins. We found markedly decreased surface expression of CXCR4 and MPL, two master regulators of HSC quiescence and niche interaction, in Sel1L-knockout HSCs. Tracking HSC and niche cells at the single cell level in vivo showed aberrant localization of Sel1L-deficient HSCs in the bone marrow niche. We hypothesize that Sel1L ERAD governs HSC quiescence and self-renewal by regulating HSC transmembrane receptor maturation and HSC-niche interaction. We will establish the physiological significance of Sel1L ERAD in the maintenance of HSCs (Aim 1), determine the significance and mechanism of the ERAD- Unfolded Protein Response (UPR) crosstalk in HSCs (Aim 2), and elucidate the mechanism and significance of Sel1L ERAD in HSC-niche interactions (Aim 3). This study will provide significant insight into the post- translational regulation of HSC quiescence, self-renewal, and niche interaction by ER protein quality control mechanisms, and further identify novel determinants of HSC fates.